1. Field of the Invention
The present invention relates to the microbiological industry, and specifically to a method for producing an L-amino acid using a bacterium of the Enterobacteriaceae family which has been modified to attenuate expression of the rspAB operon.
2. Brief Description of the Related Art
Conventionally, L-amino acids are industrially produced by fermentation methods utilizing strains of microorganisms obtained from natural sources, or mutants thereof. Typically, the microorganisms are modified to enhance production yields of L-amino acids.
Many techniques to enhance L-amino acid production yields have been reported, including transformation of microorganisms with recombinant DNA (see, for example, U.S. Pat. No. 4,278,765). Other techniques for enhancing production yields include increasing the activities of enzymes involved in amino acid biosynthesis and/or desensitizing the target enzymes of the feedback inhibition by the resulting L-amino acid (see, for example, WO 95/16042, or U.S. Pat. Nos. 4,346,170, 5,661,012, and 6,040,160).
Another way to enhance L-amino acid production yields is to attenuate expression of a gene, or several genes, involved in degradation of the target L-amino acid, genes diverting the precursors of the target L-amino acid from the L-amino acid biosynthetic pathway, genes involved in the redistribution of carbon, nitrogen, and phosphate fluxes, and genes coding for toxins etc.
When the nutrient suppy becomes insufficient, many bacteria differentiate and become resistant to environmental stresses. For Escherichia coli, this process is mediated by the sigma S subunit of RNA polymerase. Expression of sigma S is induced by homoserine lactone, a metabolite synthesized from the intermediates in threonine biosynthesis. Homoserine lactone-dependent synthesis of sigma S was prevented by overexpression of the protein RspA. The function of homoserine lactone derivatives in many cell density-dependent phenomena and the similarity of RspA to a Streptomyces ambofaciens protein suggest that synthesis of homoserine lactone may be a general signal of starvation (G. W. Huisman and R. Kolter, Science, 265:537-539 (1994)).
rspB is likely to encode a catabolic enzyme because its gene product RspB is 38% identical to threonine dehydrogenase from E. coli (G. W. Huisman and R. Kolter, Science, 265:537-539 (1994), B. D. Aronson et. al., J. Biol. Chem., 264:5226-32 (1989)).
The genes rspA and rspB form an operon. This is based on data which shows that transposon insertion into rspA abolishes rspB expression (G. W. Huisman and R. Kolter, Science, 265:537-539 (1994).
But currently, there are no reports of attenuating expression of the rspAB operon for the purpose of producing L-amino acids.